大鼠杏仁体和海马结构内CREB对应激刺激的调节作用

发布时间：2018-03-15 02:07

  本文选题：CREB　切入点：杏仁体　出处：《复旦大学》2005年博士论文　论文类型：学位论文

【摘要】：杏仁体(amygdala, AM)和海马结构(hippocampal formation, HF)是与情绪活动和学习记忆相关的重要结构,AM和HF内的转录因子cAMP反应元件结合蛋白(CREB)被证实在记忆调节过程中发挥了重要作用。CREB是一种核蛋白,是刺激诱导转录因子的主要成员,多种刺激都可导致CREB的活化,形成磷酸化的CREB(pCREB)。神经生理及神经药理学的研究表明,在突触可塑性的长时程效应阶段,CREB可调节基因转录、参与蛋白质合成,从而维持突触效率的长期持续改变。但是形态上,目前还不清楚AM、HF等脑区内的CREB究竟在哪类神经元中表达而发挥作用。为此本实验准备进一步探讨:① 应激刺激后AM和HF内pCREB表达量的变化;② pCREB在什么神经元中表达;③ pCREB的表达与NMDA受体活性的关系。本实验作了如下工作: 1.对SD大鼠强迫性游泳(FS),建立情绪性应激刺激动物模型,以正常同类大鼠为对照,用免疫细胞化学单标法、蛋白免疫印迹实验,以抗pCREB抗体观察检测pCREB免疫阳性神经元在对照组和实验组大鼠AM各亚核团和HF内的分布特征及变化规律;用免疫细胞化学双标法和免疫电镜,以抗pCREB、抗Glu和抗PV抗体,分析了表达pCREB神经元的类型。结果显示:①FS刺激后,pCREB免疫阳性细胞核在AM的外测核(LA)、基底外侧核(BL)、中央核(Ce)以及HF的齿状回(DG)和CA3区的数量明显增加。FS 1h后Ce内pCREB阳性细胞核数目最多,为1082±229个/mm~2;而LA内的pCREB阳性细胞核数目变化最大,对照组为578±106个/mm~2;而FS 15min后则为1078±338个/mm~2。在DG颗粒细胞层内,对照组的pCREB阳性细胞核数目很少,成线状散在分布;而FS后pCREB阳性细胞核的数目明显增多,成带状密布于整个DG颗粒细胞层。FS后,AM各亚核团内pCREB免疫阳性细胞核的变化趋势具有时间相关性,本实验观察到,LA内的pCREB表达量在FS后15min达到高峰,而Ce内的pCREB则在FS后1h表达量最高。免疫蛋白印迹的实验结果与上述变化密切相关,显示,FS刺激后AM各亚核和HF内pCREB总蛋白量都明显增加。②免疫细胞化学双标结果显示,在LA、BL、Ce、DG和CA3区中,呈绿色荧光的pCREB免疫阳性神经元细胞核,位于呈红色Glu免疫阳性的神经元胞质内,而未见其在
[Abstract]:Amygdala (AM) and hippocampal formation (HFF) are important structures associated with emotional activity and learning and memory. The transcription factor cAMP response element binding protein (cAMP) in HF has been proved to play an important role in memory regulation. CREB is a nuclear protein. Multiple stimuli can activate CREB and form phosphorylated CREBPCREB.Neurophysiological and neuropharmacological studies have shown that CREB can regulate gene transcription during the long term effect of synaptic plasticity. Participate in protein synthesis, thereby sustaining long-term changes in synaptic efficiency. But morphologically, It is not clear what kind of neurons CREB is expressed in the brain regions such as AMN / HF. Therefore, this study is going to further explore the changes of pCREB expression in AM and HF after the stress of 1: 1 in which neurons do the 2 pCREB express? The relationship between the expression of pCREB and the activity of NMDA receptor. 1. The animal model of emotional stress stimulation was established in SD rats by compulsive swimming. The normal rats were used as control, immunocytochemical single labeling method and Western blot were used. The distribution and changes of pCREB immunoreactive neurons in the subnuclei and HF of AM in the control group and experimental group were observed and observed by using anti pCREB antibody, and the immunocytochemical double labeling method and immunoelectron microscope were used to detect the distribution of pCREB immunoreactive neurons in the control group and the experimental group, and the anti-pCREBand anti Glu and anti PV antibodies were used. The expression of pCREB neurons was analyzed. The results showed that the number of pCREB immunoreactive nuclei in the outer nucleus of AM, the basolateral nucleus, the central nucleus, and the number of the dentate gyrus DGG) and the CA3 region of HF increased significantly. FS 1 h after stimulation, the number of pCREB in ce was significantly increased. The number of positive nuclei is the highest. The number of pCREB positive nuclei in LA was the highest, 578 卤106 / mm2 in control group and 1078 卤338 / mm-2 in DG granular cell layer 15 minutes after FS. In DG granular cell layer, the number of pCREB positive nuclei in control group was very small and distributed linearly. However, the number of pCREB positive nuclei increased significantly after FS, and the change trend of pCREB immunoreactive nuclei in the whole DG granular cell layer. FS was time-dependent. In this study, we observed that the expression of pCREB in LA reached its peak at 15 minutes after FS, while the expression of pCREB in ce was the highest at 1 hour after FS. The results of Western blot were closely related to the above changes. The results showed that the total protein content of pCREB in AM subnuclei and HF increased significantly after FS-stimulation. The results showed that in the DG and CA3 regions of BLCE-DG and CA3, the nuclei of pCREB immunoreactive neurons were green fluorescent. Located in the cytoplasm of red Glu immunoreactive neurons, but not found in the
【学位授予单位】：复旦大学
【学位级别】：博士
【学位授予年份】：2005
【分类号】：R329

【引证文献】

相关博士学位论文 前1条

1 林栋;针刺大鼠外关穴对不同脑区CREB磷酸化及相关信号转导机制研究[D];广州中医药大学;2011年

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